DNA-diagnosis of carriers of the Duchenne muscular dystrophy gene

Duchenne muscular dystrophy carrier detection has been performed by using probes XJ1.1 (intragenic probe) and probe 754 for a girl. The carrier probability was estimated by means of a computer program GenRisk combining pedigree and DNA-probe data and turned out to be 95%.     

Results of blind testing a method to detect carriers of the Duchenne muscular dystrophy gene.

We blind-tested a method that in earlier studies had shown increased leucine uptake in muscle fibers of biopsy specimens from three obligate carriers and seven of 11 putative carriers of the gene for Duchenne muscular dystrophy. Here, muscle samples obtained at biopsy in seven obligate carriers and nine control subjects from Brazil were examined in Canada without knowledge of the carrier status or serum enzyme concentrations. Leucine uptake was increased in four controls and within normal range in four carriers, a rate of false-positives and -negatives that underscores the need for blind-testing methods for detecting carriers of this disease.     

Allosteric transition of erythrocyte alkaline phosphatase from Duchenne muscular dystrophy (DMD) patients and Duchenne muscular dystrophy carriers (Homo sapiens)

1. The kinetic properties of the p-nitrophenylphosphatase (EC 3.1.3.1) from erythrocytes was investigated in DMD-patients and DMD-carriers. 2. A different allosteric behaviour in the p-nitrophenylphosphatase from DMD-patients and DMD-carriers compared to controls is supported by the following findings: (a) values of n altered in F- inhibition of (K+)-activated p-nitrophenylphosphatase with Hill coefficients -1.5, -2.2 and -3.1; (b) heterotropic effect of increased concentration of Mg2+ on F- inhibition which is reverted by K+ in DMD-carriers and in control, but not in DMD-patients. 3. Evidence is presented showing that in DMD-patients and in DMD-carriers the interaction membrane-enzyme is different from the corresponding controls.     

miRNome profiling in Duchenne muscular dystrophy; identification of asymptomatic and manifesting female carriers

Duchenne muscular dystrophy (DMD) is a fatal neuromuscular disorder that occurs due to inactivating mutations in DMD gene, leading to muscular dystrophy. Prediction of pathological complications of DMD and the identification of female carriers are important research points that aim to reduce disease burden. Herein, we describe a case of a late DMD patient and his immediate female family members, who all carry same DMD mutation and exhibited varied degrees of symptoms. In our study, we sequenced the whole miRNome in leukocytes and plasma of the family members and results were validated using real-time PCR. Our results highlighted the role of miR-409-3p, miR-424-5p, miR-144-3p as microRNAs that show correlation with the extent of severity of muscular weakness and can be used for detection of asymptomatic carriers. Cellular and circulating levels of miR-494-3p had shown significant increase in symptomatic carriers, which may indicate significant roles played by this miRNA in the onset of muscular weakness. Interestingly, circulating levels of miR-206 and miR-410-3p were significantly increased only in the severely symptomatic carrier. In conclusion, our study highlighted several miRNA species, which could be used in predicting the onset of muscle and/or neurological complications in DMD carriers.     

Human X chromosome markers and Duchenne muscular dystrophy.

Two DNA markers, a random DNA fragment 754 and the cDNA sequence encoding the gene for ornithine transcarbamylase (OTC) have been studied in kindreds segregating for Duchenne muscular dystrophy. 754 and OTC are located close physically to the mutation in the region Xp21 below the breakpoints in two Duchenne females. The genetic distance was found to be approximately 10cM between 754 and DMD (two crossovers in 26 meioses) and to be approximately 10cM between OTC and DMD (two crossovers in 26 meioses). Physical data suggest the order DMD-754-OTC. The frequency of recombination compared to physical distance between these markers and DMD suggests that there may be a hot spot of recombination. The relevance of these observations for the isolation of the DMD mutation and clinical use of these probes is discussed.     

RFLPs for Duchenne muscular dystrophy cDNA clones 9 and 10.

The complete 14-kb cDNA for the gene causing the X-linked recessive muscular dystrophy (MD) type Duchenne (DMD) and Becker (BMD) has recently been cloned and made available for deletion/duplication screening in patients. It detects 65 exon-containing nonpolymorphic HindIII fragments spread over a gene locus of about 2,000 kb. When the entire DMD cDNA is used, deletions/duplications can be found in about 65%-70% of affected patients, permitting direct carrier detection by densitometric scanning. But in cases where no deletion/duplication is detectable, RFLP analysis, specially favored within the gene, will be the method for carrier-status determination. Clones 9 and 10-1.2-kb and 0.7-kb fragments, respectively, of the 14-kb DMD cDNA--have been hybridized with human genomic DNA digested by nine different restriction enzymes. Five RFLPs, involving Asp700, PvuII, XbaI, and EcoRV sites, were detected, and Mendelian inheritance could be demonstrated. Since clones 9 and 10 are localized telomeric to the mutation-hot-spot region, their polymorphisms are thought to be very helpful as flanking markers for indirect carrier detection in families with a family history of DMD/BMD. Moreover, these RFLPs can be used for direct carrier detection or exclusion in families with patients showing a deletion/duplication in the region of p9 or p10.     

The molecular and biochemical basis of Duchenne muscular dystrophy.

Duchenne and Becker muscular dystrophy (DMD, BMD) have both been clinically recognized for over 100 years, yet throughout much of that time nothing beyond clinical evaluation and supportive care during the disease course was available to patients. The identification of the molecular basis of DMD/BMD in 1986 paved the way for extensive progress toward the understanding, diagnosis and treatment of this disease.